The N-terminal Arg2, Arg3 and Arg4 of human lactoferrin interact with sulphated molecules but not with the receptor present on Jurkat human lymphoblastic T-cells

Biochemical Journal - Tập 327 Số 3 - Trang 841-846 - 1997
Dominique Legrand1, Patrick H.C. van Berkel2, Valérie Salmon1, A. Harry VAN VEEN3, Marie‐Christine Slomianny1, H. Jan NUIJENS3, Geneviève Spik1
1Laboratoire de Chimie Biologique et Unité Mixte de Recherche no. 111 du Centre National de la Recherche Scientifique, Université des Sciences et Technologies de Lille, 59655 Villeneuve d'Ascq cedex, France
2Leiden Institute of Chemistry, Medical Biotechnology Department, Gorlaeus Laboratories, Leiden University, 2333 CA, Leiden, The Netherlands
3Pharming BV, Niels Bohrweg 11-13, 2333 CA, Leiden, The Netherlands

Tóm tắt

We previously characterized a 105 kDa receptor for human lactoferrin (hLf) on Jurkat human lymphoblastic T-cells. To delineate the role of the basic cluster Arg2-Arg3-Arg4-Arg5 of hLf in the interaction with Jurkat cells, we isolated N-terminally deleted hLf species of molecular mass 80 kDa lacking two, three or four N-terminal residues (hLf-2N, hLf-3N and hLf-4N) from native hLf that had been treated with trypsin. Native hLf bound to 102000 sites on Jurkat cells with a dissociation constant (Kd) of 70 nM. Consecutive removal of N-terminal arginine residues from hLf progressively increased the binding affinity but decreased the number of binding sites on the cells. A recombinant hLF mutant lacking the first five N-terminal residues (rhLf-5N) bound to 17000 sites with a Kd of 12 nM. The binding parameters of bovine lactoferrin (Lf) and native hLf did not significantly differ, whereas the binding parameters of murine Lf (8000 sites; Kd 30 nM) resembled those of rhLf-5N. Culture of Jurkat cells in the presence of chlorate, which inhibits sulphation, decreased the number of binding sites for both native hLf and hLf-3N but not for rhLf-5N, indicating that the hLf-binding sites include sulphated molecules. We propose that the interaction of hLf with a large number of binding sites (approx. 80000 per cell) on Jurkat cells is dependent on Arg2-Arg3-Arg4, but not on Arg5. Interaction with approx. 20000 binding sites per cell, presumably the hLf receptor, does not require the first N-terminal basic cluster of hLf. Moreover, the affinity of hLf for the latter binding site is enhanced approx. 6-fold after removal of the first basic cluster. Thus N-terminal proteolysis of hLf in vivo might serve to modulate the nature of its binding to cells and thereby its effects on cellular physiology.

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Biochemical Journal

Tập 327 Số 3

841-846

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